This application is in response to PAS-15-029 Promoting Research in Basic Neuroscience which aims to stimulate research addressing fundamental questions in basic neuroscience. Modulation of the CB1 receptor has been a research topic of great interest due to the promise of the CB1 receptor in the treatment of a range of disorders such as obesity, drug addiction, pain and inflammation. The untoward side effects associated with the use of the CB1 receptor antagonist/inverse agonist rimonabant (SR141716), including anxiety and depression, have adversely impacted the development of cannabinoid receptor antagonists and their clinical application. Recently, several classes of small molecule CB1 allosteric modulators have been discovered which offer a much needed alternative strategy to modulate CB1 signaling for therapeutic benefit. However, the specific nature of the allosteric modulation of these molecules remains unclear and there seems to be a lack of correlation between the in vitro and in vivo pharmacology. While Org27569 and PSNCBAM-1 acted as antagonists in a number of in vitro functional assays, Org27569 did not enhance or block CB1 agonist-induced effects in several animal models in mice or rats. Interestingly, we found that Org27569 dose-dependently attenuated both cue- and drug-induced reinstatement of cocaine- and methamphetamine-seeking behavior. These findings suggest CB1 allosteric modulator signaling is highly pathway and probe dependent. To better understand the signaling mechanism of CB1 allosteric modulators and develop improved molecular probes that may be pathway selective, we herein propose to design and synthesize CB1 allosteric modulators, and use an array of in vitro assays to characterize probe dependent effects on signaling, and employ these probes to investigate different signaling pathways in in vivo models to link quantifiable patterns of efficacy with in vivo phenotypic responses.